It is becoming apparent that one consequence of the hepatic effects of ethanol is increased generation of reactive oxygen intermediates. Specific objectives of this application are to evaluate the role of oxygen radicals in the mechanism by which ethanol exerts its hepatotoxic effects and if the alcoholic liver is especially sensitive to oxidative stress. Ethanol-derived NADH has been implicated in several of the metabolic actions of ethanol. Experiments will be conducted to evaluate whether NADH derived from the oxidation of ethanol can promote generation of reactive oxygen intermediates by microsomes (O2.-, H2O2, .OH, lipid peroxidation, chemiluminescence, inactivation, of added enzymes, reduction of ferric complexes). Since ferritin is likely the most important physiological source of iron, the ability of ferritin to catalyze NADH- and NADPH-dependent generation of active oxygen radicals and interact with pro-oxidants will be determined. The ability of other organelles with NAD(P)H-dependent electron transfer systems (nuclei, mitochondria, plasma membranes) to interact with iron complexes and pro-oxidants to generate reactive oxygen intermediates will be determined. After characterizing the above with control preparations, the effects of acute and chronic ethanol treatment will be assessed. ESR spectroscopy will be conducted to validate that the generated oxidant is .OH. The possible zonation of the liver with respect to oxygen radical production, anti- oxidant defense and sensitivity to oxidants will be evaluated and the effects of ethanol determined, to evaluate whether the preferential perivenous toxicity of ethanol may be related to reactive oxygen intermediates. The metabolic, acute and chronic effects of ethanol on oxygen radical generation by intact cells and in-vivo will be determined, as well as the sensitivity of hepatocytes from alcohol-treated rats to oxidants generated internally and externally. Attempts will be made to prevent some of the hepatotoxic effects of ethanol (fatty liver, glycogen depletion, mitochondrial damage) by agents which prevent or scavenge oxidizing radicals, e.g., desferrioxamine, anti-oxidants. It is hoped that these studies will provide information on the role of reactive oxygen intermediates and oxidative stress in alcohol toxicity, and be of value in attempts to prevent or ameliorate some of the toxic effects associated with alcohol abuse.